Dissertations / Theses on the topic 'Human skeletal muscle myoblast'
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Cesare, Maria Michela. "ANTIOXIDANT PROTECTION OF TUSCAN TOMATO PEEL POLYPHENOLS IN A CELLULAR MODEL OF SARCOPENIA." Doctoral thesis, Università di Siena, 2022. http://hdl.handle.net/11365/1186467.
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Background: Tomato by-products contain a great variety of biologically active substances and might represent a significant source of natural antioxidant supplements of the human diet. The preliminary studies were carried out on two ancient Tuscan tomato peel varieties, Rosso di Pitigliano (RED) and Perina a Punta della Valtiberina (PER), obtained by growing plants in normal (-Ctr) or in drought stress conditions (-Ds) present in the Regional Bank of the Germplasm of Tuscany. The variety chosen was Rosso di Pitigliano for the best beneficial effects on vascular related dysfunction. The preliminary aim of the thesis was to create an in vitro model of sarcopenia, induced by dexamethasone using human skeletal muscle myoblasts (HSMM). Sarcopenia is a disease that affects athletes who practice endurance physical activity. In these, an excessive exercise increased reactive oxygen species (ROS) levels, that, if not properly balanced by the endogenous antioxidant system, can compromise the performance of the athletes. Furthermore, in controlling muscle mass an important role is played by serine/threonine kinase and a decreased activation of the Akt-mTOR pathway by sarcopenia contributes to protein synthesis reduction. The main aim of study was to evaluate the cytoprotective properties of tomato peel polyphenols from Rosso di Pitigliano, cultivated in normal or in drought stress conditions, on an in vitro model of sarcopenia.
Methods: The antioxidant activity and total polyphenol content (TPC) were measured. The identification of bioactive compounds of several tomato peel was performed by HPLC. HUVEC were pre-treated with different TPC of RED-Ctr or RED-Ds, then stressed with H2O2. Cell viability, ROS production and CAT, SOD and GPx activities were evaluated. Permeation of antioxidant molecules contained in RED across excised rat intestine was also studied. The phenol content of both peel extracts was investigated by Ultra High-Performance Liquid Chromatography (UHPLC) analyses coupled to electrospray ionization high-resolution mass spectrometry (ESI-HR-MS). Morphological sarcopenia induction and treatment with tomato peels extracts were performed. The effector’s expression was evaluated by Real-Time PCR reactions after setting the optimal reaction conditions. Myotubes-differentiated were examined for the expression of Myosin heavy chain-2 (MYH2), Troponin T type 1 (TNNT) and Myogenin (MYOG). Furthermore, Protein kinase B (AKT1) and Forkhead Box O1 (FOXO1) mRNA expression was evaluated. Superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) activities were performed.
Results: RED-Ds tomato peel extract possessed higher TPC than compared to RED-Ctr (361.32 ± 7.204 mg vs. 152.46 ± 1.568 mg GAE/100 g fresh weight). All extracts were non-cytotoxic. Two hours pre-treatment with 5 μg GAE/mL from RED-Ctr or RED-Ds showed protection from H2O2-induced oxidative stress and significantly reduced ROS production raising SOD and CAT activity (* p < 0.05 and ** p < 0.005 vs. H2O2, respectively). The permeation of antioxidant molecules contained in RED-Ctr or RED-Ds across excised rat intestine was high with non-significant difference between the two RED types (41.9 ± 9.6% vs. 26.6 ± 7.8%).
Phenolic acids increase in the stressed tomato peel extract, while flavonoids decrease. Data shows a protective effect of 5μg GAE/ml TPC of Red DS extract on the sarcopenia. FOXO1 mRNA expression was significantly increased when cells treated with Dexa, but this expression was significantly decreased in Red Ds+Dexa (p <0.0001 vs control). AKT1 mRNA expression was increased in myotubes pre-treated with Red Ds and Dexa (p <0.0001 vs control). Myosin heavy chain 2 (MYH2), troponin T (TNNT1), miogenin (MYOG), were express in myotubes differentiated (p<0.001 vs Control). DEXA significantly reduces the antioxidant enzyme activity of SOD compared with untreated cells (p < 0.0001), but RED-Ds increased SOD activity. Conclusions: The final results show that the tomato peel extract of Rosso di Pitigliano, grown in conditions of drought stress, represents a good source of bioactive molecules, which protects the endothelium from oxidative stress even at low concentrations. Furthermore, the polyphenols from tomato peel show a cytoprotective effect in the in vitro model of sarcopenia without the use of vehicles for absorption.
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Wilson, Alyssa A. "Exploring the Role of Myoblast Fusion in Skeletal Muscle Development and Homeostasis." University of Cincinnati / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1504781294099666.
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Tallon, Mark J. "Carnosine metabolism in human skeletal muscle." Thesis, University of Chichester, 2005. http://eprints.chi.ac.uk/843/.
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Aviss, Kathryn Jane. "A synthetic biodegradable oriented scaffold for skeletal muscle tissue engineering." Thesis, University of Manchester, 2011. https://www.research.manchester.ac.uk/portal/en/theses/a-synthetic-biodegradable-oriented-scaffold-for-skeletal-muscle-tissue-engineering(baed422d-940f-4489-b180-0bed3f4fc6ee).html.
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The aim of this project was to create a novel biodegradable, synthetic scaffold that will provide the correct topographical cues for myoblast alignment and efficient differentiation into myotubes. Skeletal muscle repair after major surgery or serious burns is often overlooked leading to poor healing and consequent loss of power in movements of affected limbs. In order to overcome this problem a tissue engineered construct could be utilised as a grafting patch to encourage further regeneration and enhance possible power to the limb. Using a biodegradable polymer can provide structural support until the tissue is established, and will be excreted by the body's natural processes as it degrades. A synthetic polymer is desirable as it can reduce the risk of immunogenic responses thus reduce risk of graft rejection. For successful in vitro growth of skeletal muscle, the cells must be encouraged to arrange themselves into parallel arrays in order for efficient fusion and consequent contraction. By incorporating the correct topographical cues into the scaffold to promote contact guidance for cellular alignment this can be achieved. Electrospinning is a reliable technique which yields highly reproducible aligned fibres from the micro- to the nanoscale. This project focuses upon creating and characterising the electrospun scaffold, checking biocompatibility with myoblasts by monitoring the topography, residual solvent within the scaffold, the mechanical properties of the scaffold, and a brief investigation into the degradation profile of the electrospun fibres. The immunogenicity of the scaffold was investigated by monitoring cytokine release from macrophages. Myoblast morphology was monitored, as was the efficiency of the cells to differentiate and their potential to become contractile myofibres. Cellular adhesion to the scaffold was also looked into by measuring the expression of integrins during early and late adhesion and on substrates with different topographies. It was found that the electrospun scaffold did not contain a significant amount of residual solvent, and macrophages were not activated any more than on tissue culture plastic. Myoblasts responded to the topography of the aligned fibres by aligning along the length of the fibres, showing elongation and bi-axial cytoskeletal arrangement after just 30 minutes culture on the aligned fibres. This elongation prompted fusion and differentiation of the myoblasts to occur faster than cells which were not exposed to the aligned topography, and this global alignment was maintained in long term culture.
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Leng, Xinyan. "Roles of proteasome, arachidonic acid, and oxytocin in bovine myoblast proliferation and differentiation." Diss., Virginia Tech, 2018. http://hdl.handle.net/10919/82707.
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The overall objective of this dissertation project was to identify factors and mechanisms that control bovine myoblast proliferation, differentiation, and fusion. Three studies were conducted during this project. The objective of the first study was to determine the effect of oxytocin (OXT) on myoblast proliferation, differentiation and fusion. Treating primary bovine myoblasts in culture with 10 nM and 100 nM OXT for 24 h increased their proliferation rate by 7% (P < 0.05) and 10% (P < 0.05), respectively. Treating bovine myoblasts with either concentration of OXT for 48 h had no effect on their differentiation and fusion, as indicated by no changes in mRNA expression of selected myoblast differentiation markers and fusion index. The objective of the second study was to determine the effects of arachidonic acid (AA) and its major metabolites prostaglandin E2 (PGE2), PGF2a, and PGI2 on myoblast proliferation, differentiation and fusion. Treating myoblasts with 10 μM AA, 1 μM PGE2, 1 μM PGF2α, and 1 μM PGI2 for 24 h each increased the number of proliferating cells by 13%, 24%, 16%, and 16%, respectively, compared to the control (P < 0.05). At the same concentrations, AA, PGE2, and PGF2a stimulated myoblast differentiation and PGE2 improved myoblast fusion (P < 0.05). Treating myoblasts with AA and the cyclooxygenase (COX)-1 and COX-2 inhibitor indomethacin or the COX-2-specific inhibitor NS-398 reversed the stimulatory effect of AA on myoblast proliferation (P < 0.05). The objective of the third study was to determine the role of the proteasome in bovine myoblast differentiation and fusion. It was found that the proteasome activity increased (P < 0.05) during myoblast differentiation and fusion. Adding 5 μM lactacystin, a specific inhibitor of the proteasome, to the differentiation medium nearly completely blocked myoblast differentiation and fusion. Inhibitor of DNA-binding 1 (ID1) is known to inhibit myoblast differentiation and to be degraded by the proteasome in some cells. Both ID1 protein and mRNA expression were found to decrease during myoblast differentiation and fusion, and the decrease in ID1 protein but not ID1 mRNA was reversed (P < 0.05) by treating the cells with lactacystin. In summary, this project reveals that OXT and AA are stimulators of bovine myoblast proliferation and that AA is a stimulator of bovine myoblast differentiation. This project also indicates that the proteasome plays a positive role in bovine myoblast differentiation and fusion, and that it does so perhaps by reducing the accumulation of the ID1 protein.Ph. D.
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Renna, L. V. "MOLECULAR BASIS OF SKELETAL MUSCLE ATROPHY IN MYOTONIC DYSTROPHY." Doctoral thesis, Università degli Studi di Milano, 2015. http://hdl.handle.net/2434/333083.
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Myotonic dystrophy (DM) is an autosomal dominant multisystemic disorder characterized by a variety of multisystemic features including myotonia, muscular dystrophy, cardiac dysfunctions, cataracts and insulin-resistance. DM1 is caused by an expanded (CTG)n in the 3’ UTR of the DMPK gene, while DM2 is caused by the expansion of a (CCTG)n repeat in the intron 1 of the CNBP gene. In both forms, the mutant transcripts accumulate in nuclear foci altering the function of some alternative splicing regulators which are necessary for the physiological processing of mRNAs. However, the downstream pathways by which these RNA binding proteins cause skeletal muscle alteration are not well understood. For these reasons the aim of my PhD project was to analyze the molecular mechanisms behind DM skeletal muscle atrophy. In the first part of my PhD we have performed different studies to better define the molecular pathogenesis of DM. In particular, we have analysed the histopathological and biomolecular features of skeletal muscle biopsies from a cohort of DM1 and DM2 patients presenting different phenotypes. The results indicated that the splicing and muscle pathological alterations observed are related to the clinical DM1 and DM2 phenotype and that CUGBP1 seems to play a role only in DM1, confirming that the molecular pathomechanism of DM is more complex than the one actually suggested. These data were confirmed by the analysis of two different biopsies obtained from 5 DM2 patients that showed that morphological alterations evolve more rapidly over time than the molecular changes suggesting that the molecular mechanisms that drive to skeletal muscle atrophy are still unclear and that these features cannot be explained only by spliceopathy. For all these reasons we decided to analyse DM satellite cells activity in vitro. Satellite cells are the muscle fibre precursor cells and our data indicated that both DM1 and DM2 skeletal muscle cells have lower proliferative capability than control myoblasts. Moreover, the premature proliferative growth arrest observed in DM cells appears to be caused by an overexpression of p16 in DM1 muscle cells, while DM2 muscle cells stop dividing with telomeres shorter than controls, suggesting that in these cells the signaling involved in premature senescence depend on a telomere-driven pathway. Finally, we decided to analyze the insulin pathway which is involved in the regulation of skeletal muscle atrophy. Our data have shown that DM1 and DM2 cells exhibit a lower glucose uptake and a lower proteins activation after 10 nM insulin stimulation when compared to controls suggesting that also this pathway could play a role in the molecular mechanisms that drive skeletal muscle atrophy in DM patients. In conclusion, we have shown that the molecular mechanisms behind skeletal muscle atrophy in DM1 and DM2 patients are more complicated than that previously suggested and further analysis are necessary to understand why skeletal muscle atrophy affect mainly type 1 fibres in DM1 patients, while on the contrary it affects selectively type 2 fibres in DM2 patients.
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Adamo, Kristi Bree. "Proglycogen and macroglycogen in human skeletal muscle." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp01/MQ31807.pdf.
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Saxton, John Michael. "Exercise-induced damage to human skeletal muscle." Thesis, University of Wolverhampton, 1990. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.385185.
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Hurel, Steven J. "Insulin action in cultured human skeletal muscle." Thesis, University of Newcastle Upon Tyne, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.363891.
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Ruiz, Carlos Ariel. "Transcriptional and Post-Transcriptional Regulation of Synaptic Acetylcholinesterase in Skeletal Muscle." Scholarly Repository, 2009. http://scholarlyrepository.miami.edu/oa_dissertations/370.
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myotubesProper muscle function depends upon the fine tuning of the different molecular components of the neuromuscular junction (NMJ). Synaptic acetylcholinesterase (AChE) is responsible for rapidly terminating neurotransmission. Neuroscientists in the field have elucidated many aspects of synaptic AChE structure, function, and localization during the last 75 years. Nevertheless, how the enzyme is regulated and targeted to the NMJ is not completely understood. In skeletal muscle the synaptic AChE form derives from two separate genes encoding the catalytic and the collagenic tail (ColQ) subunits respectively. ColQ-AChE expression is regulated by muscle activity; however, how this regulation takes place remains poorly understood. We found that over or down-regulation of ColQ is sufficient to change the levels of AChE activity by promoting assembly of higher order oligomeric forms including the collagen-tailed forms. Furthermore, when peptides containing the Proline Rich Attachment Domain (PRAD), the region of ColQ that interacts with the AChE, are fed to muscle cells or cell lines expressing AChE, they are taken up by the cells and retrogradely transported to the endoplasmic reticulum (ER)/Golgi network where they induce assembly of newly synthesize AChE into tetramers. This results in an increase, as a consequence, in total cell associated AChE activity and active tetramer secretion, making synthetic PRAD peptides potential candidates for the treatment of organophosphate pesticides and nerve gas poisoning. To study the developmental regulation of ColQ-AChE we determined the levels of ColQ and ColQ mRNA in primary quail muscle cells in culture and as a function of muscle activity. Surprisingly, we found dissociation between transcription and translation of ColQ from its assembly into ColQ-AChE indicating the importance of posttranslational controls in the regulation of AChE folding and assembly. Furthermore, we found that the vast majority of the ColQ molecules in QMCs are not assembled into ColQ-AChE, suggesting that they can have alternative function(s). Finally, we found that the levels of ER molecular chaperones calnexin, calreticulin, and particularly protein disulfide isomerase are regulated by muscle activity and they correlate with the levels of ColQ-AChE. More importantly, our results suggest that newly synthesized proteins compete for chaperone assistance during the folding process.
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Stephens, Francis B. "Carnitine transport and metabolism in human skeletal muscle." Thesis, University of Nottingham, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.430645.
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Pickersgill, Laura. "Lipid-induced insulin resistance in human skeletal muscle." Thesis, University of Newcastle Upon Tyne, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.413955.
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Kennedy, Paul. "Magnetic resonance elastography studies of human skeletal muscle." Thesis, University of Edinburgh, 2016. http://hdl.handle.net/1842/25776.
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A robust, reliable method to non-invasively measure in-vivo mechanical properties of large tissue areas was unavailable until the advent of a new Magnetic Resonance Imaging (MRI) technique known as Magnetic Resonance Elastography (MRE). MRE is a phase-contrast imaging technique that enables quantification of tissue mechanical properties by capturing the motion of induced shear waves via a synchronised Motion Encoding Gradient (MEG). The complex shear modulus is determined via mathematical inversion and reported as the magnitude of the complex shear modulus, |G*|, and phase angle, φ. The work reported in this thesis focuses on the development of MRE data acquisition and analysis protocols optimised to study thigh muscle mechanical properties. The protocols are subsequently applied in healthy volunteers to study natural phenomena such as contraction and ageing and interventions such as an experimental protocol to produce Exercise Induced Muscle Damage (EIMD). Methodological advances made throughout this work include moving from 2D to 3D MRE data acquisition protocols and the application of advanced inversion software to extract muscle viscoelastic properties from the acquired MRE data. Results obtained include observation of reduced muscle stiffness in 6 elderly subjects (>80 years old) compared to 4 young subjects in the Vastus Lateralis (32%), quadriceps muscle group (22%) and entire thigh cross-section (19%), higher resting stiffness of agonist quadriceps compared to antagonist hamstrings (18%) and an increase in quadriceps stiffness (40%) during a leg raise task in 11 healthy subjects. Variability in muscle group recruitment patterns during the contraction were also observed, with the phase angle of the Vastus Intermedius (VI) increasing significantly during contraction. The final experiment involved the recruitment of 20 healthy male subjects to perform an eccentric exercise protocol designed to induce EIMD. Subjects who displayed a Maximum Voluntary Contraction (MVC) force deficit of >10% were considered to have experienced EIMD. A further severe EIMD group were defined based on the presence of hyper-intense signal on T2 weighted imaging following the protocol. The T2 hyper-intensity was found to occur exclusively in the Rectus Femoris (RF) and VI muscle groups. Increased muscle stiffness was observed in the RF muscle in subjects who experienced moderate EIMD (6%). A greater increase in RF stiffness (48%) was observed in the severe EIMD group. The severe EIMD group also displayed significantly increased VI stiffness (14%). The experiments carried out provide several novel findings which can support the development of beneficial strategies to promote both healthy ageing and rehabilitation in athletes, and potentially contribute to improving muscle performance evaluation tests which will expand the opportunities for clinical applications of muscle MRE.
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Enwere, Emeka K. "Regulation of Skeletal Muscle Formation and Regeneration by the Cellular Inhibitor of Apoptosis 1 (cIAP1) Protein." Thèse, Université d'Ottawa / University of Ottawa, 2011. http://hdl.handle.net/10393/20048.
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The inhibitor of apoptosis (IAP) proteins traditionally regulate programmed cell death by binding to and inhibiting caspases. Recent studies have uncovered a variety of alternate cellular roles for several IAP family members. The cellular inhibitor of apoptosis 1 (cIAP1) protein, for instance, regulates different axes of the NF-κB signalling pathway. Given the extensive functions of NF-κB signalling in muscle differentiation and regeneration, I asked if cIAP1 also plays critical roles in skeletal muscle myogenesis. In a primary myoblast cell-culture system, genetic and pharmacological approaches revealed that loss of cIAP1 dramatically increases the fusion of myoblasts into myotubes. NF-κB signalling occurs along a classical and an alternative pathway, both of which are highly active in cIAP1-/- myoblasts. Suppression of the alternative pathway attenuates myotube fusion in wildtype and cIAP1-/- myoblasts. Conversely, constitutive activation of the alternative pathway increases myoblast fusion in wildtype myoblasts. cIAP1-/- mice have greater muscle weight and size than wildtypes, as well as an increased number of muscle stem cells. These results identify cIAP1 as a regulator of myogenesis through its modulation of classical and alternative NF-κB signalling pathways.
Loss of the structural protein dystrophin in the mdx mouse model of Duchenne muscular dystrophy leads to chronic degeneration of skeletal muscle. The muscle pathology is strongly influenced by NF-κB signaling. Given the roles demonstrated for cIAP1 in cell culture and in vivo, I asked whether loss of cIAP1 would influence muscle pathology in the mdx mouse. To address this question, double-mutant mice were bred lacking both cIAP1 and dystrophin (cIAP1-/-;mdx). Histological analyses revealed that double-mutant mice exhibited reduced indications of damage on several measures, as compared to single-mutant (cIAP1+/+;mdx) controls. Unexpectedly, these reductions were seen in the “slow-twitch” soleus muscle but not in the “fast-twitch” extensor digitorum longus (EDL) muscle. The improvements in pathology of double-mutant solei were associated with reductions in muscle infiltration by CD68-expressing macrophages. Finally, the double-mutant mice exhibited improved endurance and resistance to damage during treadmill-running exercise. Taken together, these results suggest that loss of cIAP1, through its multiple regulatory functions, acts to improve myogenesis and increase muscle resistance to damage.
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Loiselle, Julie Jennifer. "Analysis of RBM5 and RBM10 expression throughout H9C2 skeletal and cardiac muscle cell differentiation." Thesis, Laurentian University of Sudbury, 2013. https://zone.biblio.laurentian.ca/dspace/handle/10219/2032.
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RNA Binding Motif (RBM) domain proteins RBM5 and RBM10 have been shown to influence apoptosis, cell cycle arrest and splicing in transformed cells. In this study, RBM5 and RBM10 were examined in non-transformed cells in order to gain a wider range of knowledge regarding their function. Expression of Rbm5 and Rbm10, as well as select splice variants, was examined at the mRNA and protein level throughout H9c2 skeletal and cardiac myoblast differentiation. Results suggest that Rbm5 and Rbm10 may (a) be involved in regulating cell cycle arrest and apoptosis during skeletal myoblast differentiation and (b) undergo post-transcriptional or translational regulation throughout myoblast differentiation. All in all, the expression profiles obtained in the course of this study will help to suggest a role for Rbm5 and Rbm10 in differentiation, as well as possible differentiation-specific target genes with which they may interact.
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Crowther, Gregory John. "An analysis of metabolic fluxes in contracting human skeletal muscle /." Thesis, Connect to this title online; UW restricted, 2002. http://hdl.handle.net/1773/10538.
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O'Leary, Mary Frances. "The role of adipose and skeletal muscle derived cytokines in primary human myogenesis : implications for ageing skeletal muscle." Thesis, University of Birmingham, 2018. http://etheses.bham.ac.uk//id/eprint/8089/.
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Sarcopenia is the age-related loss of skeletal muscle mass and function; inflammation is thought to be one aetiological factor in its development. Adipose tissue accumulates with advancing age and adipose-derived cytokines (adipokines) contribute to inflammaging. Skeletal muscle myogenesis is one adaptaive mechanism by which skeletal muscle mass is sustained throughout the human lifespan. The effect of the adipose inflammatory milieu on such myogenesis is unknown, as is the relative importance of its constituent adipokines to myogenesis. This work demonstrates that conditioned medium generated from obese subcutaneuous adipose tissue has a detrimental effect on in vitro primary human myogenesis. Resistin is shown to be – in part – responsible for this phenomenon and is demonstrated to inhibit myogenesis by activating the classical NFκB pathway. Resistin is further shown to be a metabolic stressor of primary human myotubes, promoting increased oxygen consumption, fatty acid oxidation and lipid accumulation. It is important to identify more avenues for the development of pharmacological interventions in sarcopenia. To that end, this thesis also demonstrates for the first time that the myokine IL-15: 1) is pro-myogenic in primary human cultures; 2) can mitigate the detrimental effects of an inflammatory environment on myogenesis; and 3) supports myogenesis at autocrine concentrations.
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Gustafsson, Thomas. "Exercise and angiogenic growth factors in human skeletal muscle /." Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-387-6/.
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Vesali, Rokhsareh Farrah. "Amino acid and protein turnover in human skeletal muscle /." Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-285-3/.
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Norton, Luke. "Calpain-10 and insulin resistance in human skeletal muscle." Thesis, University of Nottingham, 2007. http://eprints.nottingham.ac.uk/11536/.
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Variation in the calpain-10 gene has been linked to a three-fold increased risk for type 2 diabetes in Pima Indian and some European populations. Furthermore, reduced skeletal muscle expression of calpain-10 is associated with reduced insulin mediated glucose disposal and carbohydrate oxidation. The skeletal muscle specific calpain-3 plays a key role in skeletal muscle integrity and has also been linked to insulin resistance in humans and rodents. The major aims of this thesis were to 1) investigate the hypothesis that alterations in insulin sensitivity in healthy humans would lead to significant changes in the mRNA and protein expression of calpain-10 and -3, 2) investigate the effect of hyperinsulinaemia and lipid availability on calpain-10 and -3 expression, 3) further address the role of genetic variation in the calpain-10 gene on glucose utilisation in humans and finally 4) investigate the expression of calpain-10 in skeletal muscle of type 2 diabetic patients. The studies in this thesis show for the first time that insulin resistance as a result of short term fasting or high fat availability is not associated with changes in calpain-10 and -3 mRNA and protein expression, providing evidence against an adaptive role for these genes in the development of fasting- and lipid-induced insulin resistance.
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DeMenna, Jacob. "Acute Exercise Alters Promoter Methylation in Human Skeletal Muscle." Thesis, The University of Arizona, 2016. http://hdl.handle.net/10150/603565.
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A Thesis submitted to The University of Arizona College of Medicine - Phoenix in partial fulfillment of the requirements for the Degree of Doctor of Medicine.Background And Significance: Insulin resistance is an underlying disease of obesity and type 2 diabetes, which is a metabolic health crisis in the United States. Insulin resistance is caused by a combination of environmental and genetic factors. Understanding the epigenetic factors, specifically DNA methylation and how it influences the expression of genes linked to insulin resistance is of critical importance. Research Question: In this project, we set out to identify patterns of changes in DNA methylation in response to an acute exercise in healthy control subjects. Methods: Five lean (BMI = 23.6 ± 3.3 kg/m2) volunteers underwent a euglycemic hyperinsulinemic clamp with a baseline muscle biopsy and a single bout of aerobic exercise on a stationary bicycle for 48 minutes, rotating between 70 and 90% of VO2max, with a muscle biopsy taken 24 hours after completing the exercise. DNA was isolated from the baseline and 24 hours muscle biopsy, and next‐generation reduced representation bisulfite sequencing (RRBS) was performed, with analysis of the data using methylSig, and KEGG pathway analysis. Results: RRBS analysis captured 676,937 methylation sites, and of these 47,459 were differently methylated following acute exercise (P<0.05) with 4,574 sites occurring in promoter and untranslated (5’ and 3’) regions. The site with the greatest increase in methylation was within the gene NADP(+) ‐dependent malic enzyme cytosolic form (ME1) that demonstrated a significant methylation difference of +63.3%. A site in the gene for adenomatosis polyposis coli down‐regulated 1‐like (APCDD1L) was observed to have the most significant decrease in methylation by ‐65.3%. The gene with the highest incidence of differentially methylated sites was the gene for cardiomyopathy associated 5 (CMYA5) with 11 sites demonstrating a mean increase in methylation of 30.47%. The gene family with sequence similarity 176, member B protein (FAM176B) had the highest frequency of methylated sites (n=7) that were decreased in methylation with a mean decrease of ‐24.28%. KEGG pathway analysis was performed, which revealed significant (P<0.05) increases in methylation in the pathways of Wnt signaling, Heterotrimeric G‐protein signaling ‐Gi alpha and Gs alpha mediated, Cadherin signaling, Melanogenesis, Axon Guidance, and Neuroactive ligand‐receptor interaction. Significantly 4 enriched pathways with decreased methylation post exercise demonstrated one pathway, the Calcium signaling pathway. Conclusion: Our data demonstrates that a single bout of exercise can alter the DNA methylation pattern in skeletal muscle. Changes were observed in genes related to metabolic pathways, supporting previously published findings of changes in mRNA and proteins involved in metabolism following exercise. Future work is warranted with obese and type 2 diabetic participants to explore the differences in response to exercise between these groups.
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Kosek, David J. "Aging differences in mechanisms of human skeletal muscle hypertrophy." Thesis, Birmingham, Ala. : University of Alabama at Birmingham, 2007. https://www.mhsl.uab.edu/dt/2009r/kosek.pdf.
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BACI, DENISA. "Human induced pluripotent stem cells for skeletal muscle diseases." Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2014. http://hdl.handle.net/2108/201887.
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Regenerative medicine along with tissue engineering represent two closely related fields leading promising advances for the treatment of numerous musculoskeletal diseases and injuries. Nevertheless, new efforts are urgently needed to design a successful therapeutic approach for muscular disorders, aiming at identifying a functional stem cell population and biomaterial scaffolds in which cells and growth factors could be embedded. In this context, recent studies have suggested that reprogramming of somatic cells by defined transcription factors into induced pluripotent stem cells (iPS), as source for generating autologous muscle progenitor cells (MPs), overcomes several limitations related to adult myoblast therapy. The prospect of an unlimited cell source combined with properties such as a more proliferative capacity in vitro, suggesting a better regenerative capacity in vivo models, indicates that iPS could be a promising candidate for stem cell therapy to regenerate skeletal muscle. iPS have been shown to retain specific features that are remnants of epigenomes and transcriptomes of the donor tissue termed ‘epigenetic memory’. Given to these findings, during the first part of the present study, we generated iPS derived from skin fibroblasts and pericytes (known to have a remarkable myogenic capacity) from the same donor to determine whether the epigenetic memory could influence iPS properties, preferentially generating cells similar of the donor somatic cell type. Until now, different approaches have been reported to generate MPs from iPS. So far, these methods present limitations such as low efficiency/reproducibility and usually involve cell sorting for enrichment or forced expression of skeletal master genes risking undesired genetic recombination. Recently, substantial interest is mounting regarding extracellular vesicles (EVs) and their involvement in many cellular processes, including myogenesis. We explored the possibility to use EVs as "physiological liposomes" enriched with myogenic factors to trigger skeletal myogenesis. To this end, during the second part of the study we developed a new transgenic-free approach to obtain transplantable MPs by means of defined factors and extracellular vesicles (EVs) secreted from differentiated mouse skeletal myoblasts. We established a novel, robust stepwise protocol by treating iPS with a WNT agonist, CHIR 99021 and myotubes-derived EVs. Thus, this method has two main advantages: (i) studying molecular mechanisms of myogenesis which is overpassed in case of genetic manipulation; (ii) muscle progenitors are not terminally differentiated, and therefore have a better repair potential following transplantation. One of the major hurdles of stem cell therapy for skeletal muscle regeneration is the massive death following transplantation. Biomaterials exhibit immune protection properties and would ensure an artificial microenvironment which permits them to interact with host cells and exert their therapeutic benefits. With the purpose of a better engraftment, we employed Poly (ethylene glycol) (PEG) -fibrinogen hydrogel (PF) as cell carrier for skeletal muscle regeneration. When transplanted in a αsarcoglycan knockout/severe combined immunodeficiency beige (α-SGKO/SCIDbg) mice, PF-embedded myogenic progenitor cells exhibited stable long-term engraftment and participated in muscle regeneration by fusing with existing muscle fibers. Importantly, no teratoma and no abnormal structure were detected in the muscles transplanted with MPs Finally, our finding and differentiation system provide an effective method that facilitates further utilization of iPS .
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Axelson, Hans. "Muscle Thixotropy : Implications for Human Motor Control." Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis : Univ.-bibl. [distributör], 2005. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-5791.
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Elmubarak, M. H. "Effects of denervation on postnatal differentiation of rat skeletal muscle." Thesis, University of Bristol, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.375022.
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Ding, Ran. "Construction of functional artificial skeletal muscle tissue by regulation of cell-substrate interaction using myogenic C2C12 cells." Kyoto University, 2020. http://hdl.handle.net/2433/253516.
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Kyoto University (京都大学)0048
新制・課程博士
博士(人間・環境学)
甲第22671号
人博第957号
新制||人||227(附属図書館)
2020||人博||957(吉田南総合図書館)
京都大学大学院人間・環境学研究科相関環境学専攻
(主査)教授 川本 卓男, 教授 宮下 英明, 教授 高田 穣
学位規則第4条第1項該当
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Zhang, Hong. "Regulation of Skeletal Muscle Development And Differentiation by Ski." Case Western Reserve University School of Graduate Studies / OhioLINK, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=case1226938149.
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Shannon, C. E. "Skeletal muscle carnitine metabolism during intense exercise in human volunteers." Thesis, University of Nottingham, 2016. http://eprints.nottingham.ac.uk/28203/.
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Increasing skeletal muscle carnitine content enhances PDC flux during 30 minutes of continuous exercise at 80% Wmax, reducing reliance on non-mitochondrial ATP production and improving work output. These studies in healthy volunteers evaluated a carnitine feeding strategy that did not rely on the high carbohydrate load previously used, then investigated whether manipulating muscle carnitine could alter the adaptations to a period of submaximal high-intensity intermittent training (HIT). The rate of orally ingested 2H3-carnitine uptake into skeletal muscle was directly quantified for the first time in vivo and increased 5-fold following ingestion of an 80g carbohydrate formulation. This positive forearm carnitine balance was entirely blunted when the carbohydrate load was supplemented with 40g of whey protein, suggesting a novel antagonisation of insulin-stimulated muscle carnitine transport by amino acids. Skeletal muscle biopsy sampling demonstrated minimal acetylcarnitine accumulation and non-mitochondrial ATP production during single-leg knee extension at 85% Wmax, suggesting that PDC flux does not limit oxidative ATP production under these conditions. Conversely, PDC flux declined over repeated bouts of cycling at 100% Wmax, as evidenced by greater non-mitochondrial ATP production in the face of similar acetylcarnitine accumulation. This suggested that muscle carnitine availability could influence oxidative ATP delivery during submaximal HIT. Manipulation of muscle carnitine content by daily carnitine/carbohydrate feeding elevated free carnitine availability and maintained PDC flux during repeated bouts of intense exercise. However, profound improvements in oxidative ATP delivery in response to HIT eclipsed any effect of this carnitine-mediated increase in PDC flux on non-mitochondrial ATP production and indeed, carnitine supplementation did not potentiate any increases in exercise capacity above submaximal HIT alone. These novel data advance our understanding of muscle carnitine transport and the interplay between carnitine metabolism, PDC flux and non-mitochondrial ATP production during intense exercise, having important implications for the development of nutritional and exercise prescription strategies to enhance human performance and health.
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Symonds, James Matthew. "Expression of cadherins in human lymphocytes and skeletal muscle cells." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp03/MQ50890.pdf.
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Hollidge-Horvat, Melanie G. "The influence of extracellular pH on human skeletal muscle metabolism." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp03/NQ66213.pdf.
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Symonds, James Matthew. "Expression of cadherins in human lymphocytes and skeletal muscle cells." Thesis, McGill University, 1998. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=21649.
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Cadherins are a family of membrane glycoproteins involved in calcium-dependent cell-cell adhesion. The expression of cadherins in two human cell types, lymphocytes and skeletal myoblasts, was examined. Human peripheral lymphoid tissue lymphocytes were analysed for N-cadherin expression. A specific population of B lymphocytes was shown to express N-cadherin. Further, N-cadherin was shown to be expressed in a distinct pattern within the germinal centers of lymph nodes by immunohistochemical techniques. N-cadherin was also detected in a B cell population from human bone marrow and peripheral blood lymphocytes from B-cell chronic lymphocytic leukemia patients. These results suggest that N-cadherin may play a role in the development, differentiation and cell survival of B lymphocytes. Human myogenic cells were studied for cadherin expression before and after the induction of myoblast fusion. Polymerase chain reaction identified two cadherins, N-cadherin and OB-cadherin, in human myoblasts. These two cadherins were shown to be developmentally regulated during myogenesis. Collectively, these studies point to a role for cadherins in human myogenesis and lymphocyte differentiation.
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Gray, Stuart R. "Temperature and in vivo human skeletal muscle function and metabolism." Thesis, University of Strathclyde, 2007. http://oleg.lib.strath.ac.uk:80/R/?func=dbin-jump-full&object_id=21683.
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Increasing the temperature of the exercising muscle, passively or actively, leads to alterations in the contractile properties of the muscle, importantly an increase in power output. There is limited information, however, regarding the metabolic changes, if any, occurring within the muscle at higher temperatures and how these are related to the contractile changes occurring within the muscle and how such changes may, or may not, affect the efficiency of the working muscles. The greater power output produced during maximal sprint cycling, after a passive increase in Tm, was associated with an increase in the rate of anaerobic ATP turnover and muscle fibre conduction velocity. Further investigation revealed that this greater anaerobic ATP turnover within the muscle was the result of a greater activity of type HA fibres in the cadence range of 160-180 revs. min⁻¹. When the external power output of the muscle remains constant during more prolonged cycling exercise, performed at 60 revs. min⁻¹, there was also a greater rate of anaerobic ATP turnover in the first 2 min of exercise, with no differences in the remainder of exercise after passive elevation of Tm. There were no differences in the aerobic energy contribution or the kinetics of the V0₂ response between T. conditions. These changes led to a decrease in mechanical efficiency in the first 2 min of exercise, which was associated with a tendency for a greater PCr degradation in type I fibres. When T. was elevated via prior intense exercise there was decrease in mechanical efficiency, during 6 min of heavy exercise, at both 60 and 120 revs. min⁻¹. There was also a greater "absolute" primary amplitude and decrease in the slow component after prior exercise, with the response being greater at 120 revs. min⁻¹. The present research has demonstrated that whilst an increase in T. leads to a greater power output, during maximal exercise, mechanical efficiency is reduced as exercise progresses beyond a few seconds. Furthermore, at faster pedal rates T. affects type IIA fibres whilst at slower pedal rates (60 revs. min⁻¹) there appears to be a preferential effect on type I fibres, highlighting the velocity specific effect of Tm.
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Child, R. B. "Exercise and free radical induced damage to human skeletal muscle." Thesis, University of Wolverhampton, 1997. http://hdl.handle.net/2436/96616.
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Parker, Dawn Fiona. "Factors controlling the development and strength of human skeletal muscle." Thesis, University College London (University of London), 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.244016.
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Clark, Juliette A. "Subcellular distribution of lipid metabolising enzymes in human skeletal muscle." Thesis, University of Birmingham, 2012. http://etheses.bham.ac.uk//id/eprint/3378/.
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In obesity, lipids stored in muscle as lipid droplets (LDs) lead to accumulation of fatty acid (FA) metabolites and insulin resistance. This research involves development of immunofluorescence microscopy methods to generate novel information on the subcellular content and distribution of key enzymes that play a role in the underlying mechanisms. Chapters 3 and 4 describe visualisation of two lipid synthesising enzymes. Both are more abundant in type I muscle fibres. Chapter 5 reveals no differences between these enzymes in non obese and obese elderly women. Chapter 6 reveals that a key lipolytic enzyme (ATGL) has a higher content in type I fibres, but its activator does not. Chapter 7 describes visualisation of SNAP23 and reveals a high content at the plasma membrane and mitochondria and low content in LDs. Chapter 8 fails to observe a difference between obese and non obese elderly women in plasma membrane SNAP23, and therefore fails to confirm the hypothesis that LDs hijack SNAP23. However, obese women have less SNAP23 in mitochondria and this may limit FA oxidation. In conclusion this thesis describes several novel mechanisms by which obesity leads to accumulation of FA metabolites and insulin resistance. The developed methods will be a valuable novel tool for future diabetes research.
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Johnson, Andrew William. "Metabolic control of energetics in human heart and skeletal muscle." Thesis, University of Oxford, 2012. http://ora.ox.ac.uk/objects/uuid:82c0dce6-a162-4c08-b061-3ea7f2e35134.
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Myocardial and skeletal muscle high energy phosphate metabolism is abnormal in heart failure, but the pathophysiology is not understood. Plasma non-esterified fatty acids (NEFA) increase in heart failure due to increased sympathetic drive, and regulate the transcription of mitochondrial uncoupling protein-3 (UCP3), through peroxisome proliferator-activated receptor-α. The aim of the work in this thesis was to determine whether cardiac PCr/ATP ratios and skeletal muscle PCr kinetics during exercise were related to cardiac and skeletal muscle UCP3 levels respectively, thus providing a mechanism for the apparent mitochondrial dysfunction observed in heart failure. Patients having cardiac surgery underwent pre-operative testing, including cardiac and gastrocnemius 31P magnetic resonance spectroscopy. Intra-operatively, ventricular, atrial and skeletal muscle biopsies were taken for measurement of mitochondrial protein levels by immunoblotting, along with mitochondrial function by tissue respiration rates. Fasting plasma NEFA concentrations increased in patients with ventricular dysfunction and with New York Heart Association (NYHA) class. Ventricular UCP3 levels increased and cardiac PCr/ATP decreased with NYHA class, however, demonstrated no relationship to each other. In skeletal muscle, maximal rates of oxidative ATP synthesis (Qmax) related to functional capacity. Skeletal muscle UCP3 levels increased with NYHA class but were unrelated to skeletal muscle Qmax. Tissue respiration experiments revealed no relationship between ventricular function and indices of mitochondrial coupling, furthermore, indices of mitochondrial coupling were unrelated to tissue UCP3 levels. No evidence was found to support mitochondrial uncoupling, mediated through UCP3, as a cause of the abnormalities in cardiac and skeletal muscle high energy phosphate metabolism.
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Malik, Z. A. "Proteomic analysis of diurnal variation in human skeletal muscle performance." Thesis, Liverpool John Moores University, 2015. http://researchonline.ljmu.ac.uk/4511/.
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Phenotyping of human muscle based on its profile of myosin heavy chain isoforms is commonly used to help understand changes in muscle function. However, in many instances, measureable changes in force output or contractility occur in the absence of any change in myosin heavy chain profile. Therefore, more sophisticated analysis is required. Proteomic techniques including 2-dimensional gel electrophoresis, high- performance liquid chromatography and peptide mass spectrometry can be used to investigate changes in the abundance of hundreds of proteins simultaneously. To date, such techniques have not been used to specifically characterise the human myofibrillar proteome, or study how the myofibrillar proteome relates to muscle outputs such as peak isometric force or the velocity of contraction. This thesis presents a series of studies that develop proteomic techniques for the analysis of myofibrillar proteins as well as optimisation of techniques for measuring the range of muscle output from isometric through to velocity maximum of the human knee extensor muscles in vivo. After optimisation, the proteomic and muscle function measurement were employed to study diurnal variation. Time-of-day differences in sports performance and muscle function are widely reported, and typically, performance is ~10 % greater in the evening compared to the morning. This is consistent with our result in Chapter 3; we investigated this chapter by conducting a battery of muscle performance tests in a population of well-familiarised participants. Our data show that RFD exhibits the greatest diurnal variation (18 %) followed by isometric force (10.2 %). The diurnal variation in IKD data was less robust (range 8.1 - 9.8 %), which may have been due to the lesser precision of this technique compared to MVC and RFD. Therefore MVC and RFD were used in the final study. In final study, this thesis reports significantly (P<0.05) greater peak isometric force (11 %) and rate of force development (16 %) of knee extensor muscles of young strength-trained males in the evening compared to morning. Proteomic analysis of biopsy samples of the vastus lateralis profiled more than 100 myofibrillar protein species and detected 8 significant differences in protein abundance between morning and evening samples. The greatest difference was in the abundance of the slow isoform of myosin binding protein C (MyBPC1), which is known to modulate the activity of actin-bound myosin ATPases. MyBPC1 was resolved to 6 species; therefore the difference in abundance of one species reported here likely represents a change in post-translational modification. Therefore, this thesis provides associational evidence that post-translational modification of MyBPC1 contributes to the diurnal variation in muscle function.
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Fitzpatrick, Elizabeth. "Analysis of human skeletal muscle autoantibodies in myasthenia gravis patients /." The Ohio State University, 1990. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487678444257981.
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Martucci, Morena <1983>. "Aging in human liver and skeletal muscle: studies on proteasomes." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amsdottorato.unibo.it/6292/1/Martucci_Morena_tesi.pdf.
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Aging is a complex phenomenon that affects organs and tissues at a different rate. With advancing age, the skeletal muscle undergoes a progressive loss of mass and strength, a process known as sarcopenia that leads to a decreased mobility and increased risk of falls and invalidity. On the other side, another organ such as the liver that is endowed with a peculiar regenerative capacity seems to be only marginally affected by aging. Accordingly, clinical data indicate that liver transplantation from aged subjects has, in specific conditions, function and duration comparable to those achievable with grafts of liver from young donors. The molecular mechanisms involved in these peculiar aging patterns are still largely unknown, but it is conceivable that protein degradation machineries might play an important role, as they are responsible for the maintenance of cellular homeostasis. Indeed, it has been suggested that alteration of proteostasis may contribute to the onset and progression of several age-related pathological conditions, including skeletal muscle wasting and sarcopenia, as well as to the aging phenotypes.
The ubiquitin-proteasome system (UPS) is one of the most important cellular pathways for intracellular degradation of short-lived as well as damaged proteins. To date, studies on the age-related modifications of proteasomes in liver and skeletal muscle were performed prevalently in rodents, with controversial results, while only preliminary observations have been obtained in human liver and skeletal muscle. In this scenario, we want to investigate and characterize in humans the age-related modifications of proteasomes of these two different organs.
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Martucci, Morena <1983>. "Aging in human liver and skeletal muscle: studies on proteasomes." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amsdottorato.unibo.it/6292/.
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Aging is a complex phenomenon that affects organs and tissues at a different rate. With advancing age, the skeletal muscle undergoes a progressive loss of mass and strength, a process known as sarcopenia that leads to a decreased mobility and increased risk of falls and invalidity. On the other side, another organ such as the liver that is endowed with a peculiar regenerative capacity seems to be only marginally affected by aging. Accordingly, clinical data indicate that liver transplantation from aged subjects has, in specific conditions, function and duration comparable to those achievable with grafts of liver from young donors. The molecular mechanisms involved in these peculiar aging patterns are still largely unknown, but it is conceivable that protein degradation machineries might play an important role, as they are responsible for the maintenance of cellular homeostasis. Indeed, it has been suggested that alteration of proteostasis may contribute to the onset and progression of several age-related pathological conditions, including skeletal muscle wasting and sarcopenia, as well as to the aging phenotypes.
The ubiquitin-proteasome system (UPS) is one of the most important cellular pathways for intracellular degradation of short-lived as well as damaged proteins. To date, studies on the age-related modifications of proteasomes in liver and skeletal muscle were performed prevalently in rodents, with controversial results, while only preliminary observations have been obtained in human liver and skeletal muscle. In this scenario, we want to investigate and characterize in humans the age-related modifications of proteasomes of these two different organs.
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Kanaan, Georges. "Mitochondrial Dysfunction: From Mouse Myotubes to Human Cardiomyocytes." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/37582.
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Mitochondrial dysfunction is a common feature in a wide range of disorders and diseases from obesity, diabetes, cancer to cardiovascular diseases. The overall goal of my doctoral research has been to investigate mitochondrial metabolic dysfunction in skeletal and cardiac muscles in the context of chronic disease development.
Perinatal nutrition is well known to affect risk for insulin resistance, obesity, and cardiovascular disease during adulthood. The underlying mechanisms however, are poorly understood. Previous research from our lab showed that the in utero maternal undernutrition mouse model is one in which skeletal and cardiac muscle physiology and metabolism is impaired. Here we used this model to study the impact of in utero undernutrition on offspring skeletal primary muscle cells and to determine if there is a cell autonomous phenotype. Metabolic analyses using extracellular flux technologies revealed a shift from oxidative to glycolytic metabolism in primary myotubes. Gene expression profiling identified significant changes in mRNA expression, including an upregulation of cell stress and OXPHOS genes and a downregulation of cell division genes. However, there were no changes in levels of marker proteins for mitochondrial oxidative phosphorylation (OXPHOS). Findings are consistent with the conclusion that susceptibility to metabolic disease in adulthood can be caused at least in part by muscle defects that are programmed in utero and mediated by impaired mitochondrial function.
In my second project, the effects of the absence of glutaredoxin-2 (Grx2) on redox homeostasis and on mitochondrial dynamics and energetics in cardiac muscle from mice were investigated. Previous work in our lab established that Grx2-deficient mice exhibit fibrotic cardiac hypertrophy, and hypertension, and that complex I of OXPHOS is defective in isolated mitochondria. Here we studied the role of Grx2 in the control of mitochondrial structure and function in intact cells and tissue, as well as the role of GRX2 in human heart disease. We demonstrated that the absence of Grx2 impacts mitochondrial fusion, ultrastructure and energetics in mouse primary cardiomyocytes and cardiac tissue and that provision of the glutathione precursor, N-acetylcysteine (NAC) did not restore glutathione redox or prevent impairments. Furthermore we used data from the human Genotype-Tissue Expression consortium to show that low GRX2 expression is associated with increased fibrosis, hypertrophy, and infarct in the left ventricle. Altogether, our results indicate that GRX2 plays a major role in cardiac mitochondrial structure and function, and protects against left ventricle pathologies in humans.
In my third project, we collaborated with cardiac surgeon, Dr. Calum Redpath, of the Ottawa Heart Institute to study atrial mitochondrial metabolism in atrial fibrillation patients with and without type 2 diabetes (T2DM). T2DM is a major risk factor for atrial fibrillation, but the causes are poorly understood. Atrial appendages from coronary artery bypass graft surgery were collected and analyzed. We showed an impaired complex I respiration in diabetic patients with atrial fibrillation compared to diabetic patients without atrial fibrillation. In addition, and for the first time in atrial fibrillation patients, mitochondrial supercomplexes were studied; results showed no differences in the assembly of the “traditional” complexes but a decrease in the formation of “high oligomeric” complexes. A strong trend for increased protein oxidation was also observed. There were no changes in markers for OXPHOS protein levels. Overall findings reveal novel aspects of mitochondrial dysfunction in atrial fibrillation and diabetes in humans.
Overall, our results reveal that in utero undernutrition affects the programming of skeletal muscle primary cells, thereby increasing susceptibility to metabolic diseases. In addition, we show that GRX2 impacts cardiac mitochondrial dynamics and energetics in both mice and humans. Finally, we show impaired mitochondrial function and supercomplex assembly in humans with atrial fibrillation and T2DM. Ultimately, understanding the mechanisms causing mitochondrial dysfunction in muscle tissues during chronic disease development will increase our capacity to identify effective prevention and treatment strategies.
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Neji, Radhouène. "Diffusion Tensor Imaging of the Human Skeletal Muscle : Contributions and Applications." Phd thesis, Ecole Centrale Paris, 2010. http://tel.archives-ouvertes.fr/tel-00504678.
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Cette thèse propose des techniques pour le traitement d'images IRM de diffusion. Les méthodes proposées concernent l'estimation et la régularisation, le groupement et la segmentation ainsi que le recalage. Le cadre variationnel proposé dans cette thèse pour l'estimation d'un champ de tenseurs de diffusion à partir d'observations bruitées exploite le fait que les données de diffusion représentent des populations de fibres et que chaque tenseur peut être reconstruit à partir d'une combinaison pondérée de tenseurs dans son voisinage. La méthode de segmentation traite aussi bien les voxels que les fibres. Elle est basée sur l'utilisation de noyaux défini-positifs sur des probabilités gaussiennes de diffusion afin de modéliser la similarité entre tenseurs et les interactions spatiales. Ceci permet de définir des métriques entre fibres qui combinent les informations de localisation spatiale et de tenseurs de diffusion. Plusieurs approches de groupement peuvent être appliquées par la suite pour segmenter des champs de tenseurs et des trajectoires de fibres. Un cadre de groupement supervisé est proposé pour étendre cette technique. L'algorithme de recalage utilise les noyaux sur probabilités pour recaler une image source et une image cible. La régularité de la déformation est évaluée en utilisant la distortion induite sur les distances entre probabilités spatialement voisines. La minimisation de la fonctionnelle de recalage est faite dans un cadre discret. La validation expérimentale est faite sur des images du muscle du mollet pour des sujets sains et pour des patients atteints de myopathies. Les résultats des techniques développées dans cette thèse sont encourageants.
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Lindström, Mona. "Satellite cells in human skeletal muscle : molecular identification quantification and function." Doctoral thesis, Umeå universitet, Anatomi, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-29817.
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Skeletal muscle satellite cells located between the plasma membrane and the basal lamina of muscle fibres, could for many years, only be studied in situ by electron microscopy. The introduction of immunohistochemistry and the discovery of molecular markers of satellite cells then made them accessible for light microscopic studies and a wealth of information is today available. Satellite cells are myogenic stem cells that can be activated from a quiescent state to proliferate for self-renewal or differentiate into myogenic cells. The satellite cells are involved in muscle growth during fetal and postnatal development and play a key role in repair and regeneration of damaged muscle fibres. The satellite cells are also essential for muscle fibre hypertrophy and maintenance of muscle mass in the adult. When the present thesis was initiated, studies on satellite cells in human skeletal muscle relied on the neuronal cell adhesion molecule (NCAM) as a marker for satellite cell identification. The results from different studies varied markedly. Therefore the aims of the present thesis were i) to develop a highly reliable method using light microscopy for satellite cell identification and quantification in biopsies of human skeletal muscle in normal and pathological conditions. A molecular marker for the myofibre basal lamina or plasma membrane to enhance the reliability of myonuclei and satellite cell identification were to be included. Furthermore unbiased morphometric methods should be used in the quantification process. ii) to evaluate which molecular markers which had been described for satellite cell and stem cell identification in different cell states (quiescence, activated or differentiated) are the most useful for studies on human skeletal muscle. iii) to further explore the function and heterogeneity of satellite cells with respect to different markers in human skeletal muscle by studying the effects of strength-training, intake of anabolic substances and pathological conditions. A new immunofluorescence method was developed where in the same tissue section, two satellite cell markers, the basal lamina and nuclei were monitored. From the evaluation of different markers it was found that both NCAM and Pax7 identified the majority of satellite cells but that both markers were needed for reliable identification. The members of the myogenic regulatory family were evaluated and by using the new method MyoD and myogenin were found to be useful markers to identify activated and differentiated satellite cells. Upon re-examination of biopsies from power-lifters, power-lifters using anabolic substances and untrained subjects it was observed that the new results on satellite cell frequency were significantly different from those obtained when using staining for NCAM and nuclei alone. In addition three subtypes of satellite cells (94.4% NCAM+/Pax7+, 4.2% NCAM+/Pax7– and 1.4% NCAM–/Pax7+) were observed. Thus the multiple marker method gave more information about satellite cells heterogeneity in human muscle and we propose that this is more reliable than previous methods. Low numbers of MyoD or myogenin stained satellite cells were observed in both untrained and strength trained subjects. Other markers such as DLK1/FA1, a member of the EGF-like family and c-Met, the receptor for hepatocyte growth factor showed that satellite cell heterogeneity in human muscle is far greater than previously shown. Furthermore, new evidence is presented for so called fibre splitting observed in hypertrophic muscle fibres to be due to defect regeneration of partially damaged fibres.
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LeBlanc, Paul-Jean Heigenhauser George. "Dynamic and stable regulation of pyruvate dehydrogenase in human skeletal muscle /." [Hamilton, Ont.] : McMaster University, 2004.
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Karlsson, Håkan K. R. "Insulin signaling and glucose transport in insulin resistant human skeletal muscle /." Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-469-4/.
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Quisth, Veronica. "Studies on the regulation of human skeletal muscle lipolysis in vivo /." Stockholm, 2004. http://diss.kib.ki.se/2004/91-7140-167-9/.
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Downey, Jennifer. "Identification and isolation of multipotent stromal cells from human skeletal muscle." Mémoire, Université de Sherbrooke, 2013. http://hdl.handle.net/11143/6296.
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Abstract: Human skeletal muscle is an essential source of various cellular progenitors with potential therapeutic perspectives. Muscle-resident mesenchymal stromal cells (mrMSCs) are thought to be involved in the development of several regenerative disorders such as fatty degeneration, heterotopic ossification and fibrosis. Identifying the cell population responsible for these pathologies will help better understand the underlying mechanisms and lead to more efficient treatment. We first developed an isolation method and culture conditions for the proliferation and maintenance of the adherent fraction of human skeletal muscle derived cells. To further enrich the cell population as multipotent progenitors, we used fluorescent-activated cell sorting (FACS) and known mesenchymal stromal cell (MSC) markers. The enriched cell populations obtained were tested for their multipotent capabilities towards the osteogenic, adipogenic and chondrogenic lineages. The CD73 + CD 105+ CD90- subset of human skeletal muscle adherent cells displayed robust multipotence to all three lineages under the appropriate differentiation conditions. Clonal differentiation assays confirmed that all three lineages stem from a single multipotent progenitor. Furthermore, this cell subset was able to differentiate into brown adipocyte-like cells, expressing UCP1 at the RNA and protein levels following prolonged stimulation with rosiglitazone (ROS). This result suggests that this cell subset could also represent a human cell model for brown adipogenesis. The cell isolation and enrichment method presented in this thesis represent a novel technique to obtain human mrMSCs. This method holds great promise for future clinical applications with the enriched cell populations since they are expanded in a defined medium, which supports inter-laboratory reproducibility. Furthermore, the phenotypic markers chosen for the FACS isolation are well conserved amongst donors in the proposed conditions, limiting donor-to-donor variability.||Résumé: Le muscle squelettique humain est une source essentielle de cellules progénitrices ayant plusieurs applications thérapeutiques potentielles. Les cellules stromales mésenchymateuses du muscle squelettique humain (hmrMSCs) semblent être impliquées dans des pathologies telles l’ossification hétérotopique, la dégénérescence graisseuse et la fibrose. L’identification de la population cellulaire à l’origine de ces pathologies permettrait de mieux comprendre les mécanismes derrières celles-ci et aiderait à la création de traitements plus efficaces. Nous avons d’abord mis au point une méthode d'isolement et déterminer des conditions de culture pour la prolifération et le maintien en culture de la fraction cellulaire adhérente dérivée du muscle squelettique humain. Par le biais de la cytométrie en flux et des marqueurs connus des cellules stromales mésenchymateuses (MSC), nous avons pu enrichir les cellules stromales multipotentes. Le potentiel ostéogénique, adipogénique et chondrogénique des populations cellulaires enrichies a été évalué par des essais de différenciation. La sous-population de cellules CD73[indice supérieur +]CD105[indice supérieur +]CD90[indice supérieur -] a montré une multipotence robuste sur les trois lignées étudiées. Des essais de différenciation clonale ont confirmés que les trois lignées obtenues proviennent tous d’un progéniteur multipotent commun. De plus, cette sous-population cellulaire avait la capacité de se différencier en cellule de gras brun, démontrée par une expression élevée d’UCP1 au niveau génique et protéique suivant une stimulation continue avec le rosiglitazone (ROS). Ce résultat suggère que cette sous-population cellulaire pourrait également représenter un modèle pour l’adipogenèse vers le gras brun. La méthode d’enrichissement présentée représente une nouvelle technique afin d’obtenir des hmrMSCs. Elle semble prometteuse pour de futures applications cliniques employant ces cellules, étant donné qu’elles sont amplifiées dans un milieu défini permettant une reproductibilité interlaboratoire. De plus, les marqueurs de phénotype choisis pour l’enrichissement par cytométrie en flux sont bien conservés entre individus, limitant la variabilité inter-donneur.[symboles non conformes]
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McGregor, Robin A. "Skeletal muscle microRNA's in human cancer cachexia and type 2 diabetes." Thesis, Heriot-Watt University, 2009. http://hdl.handle.net/10399/2308.
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MicroRNAs are powerful post-transcriptional regulators of gene expression and are biomarkers of chronic diseases such as cancer. This thesis explores the role of microRNAs in human cancer cachexia and Type 2 diabetes. MicroRNA expression was measured in skeletal muscle biopsies using RT-qPCR. In pancreatic cancer cachexia patients, expression of microRNA-1, microRNA-133a, microRNA-133b and microRNA-206 was negatively related to weight loss. In Type 2 diabetes skeletal muscle, microRNA-133a and microRNA-206 expression was down-regulated, but there was no evidence of altered microRNA transcription or processing and target expression was unchanged. Importantly, microRNA-133a expression predicted fasting glucose, glucose tolerance and insulin resistance, and therefore may be a biomarker of Type 2 diabetes. Experimental validation of microRNA arrays was unsuccessful in identifying further novel cancer cachexia and Type 2 diabetes microRNA biomarkers. MicroRNA knockdown validated CDC42 and PTBP1 as microRNA-133a targets in myoblasts. In addition, muscle microRNA expression may be regulated by insulin and TNFa. In conclusion, microRNA-133a may be a skeletal muscle biomarker of Type 2 diabetes and cancer cachexia, microRNA-133a responds to extracellular insulin and TNFa, but it remains to be established whether microRNA-133a contributes to cancer cachexia or Type 2 diabetes pathogenesis.
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Hart, C. "The effect of critical limb ischaemia on adult human skeletal muscle." Thesis, University College London (University of London), 2013. http://discovery.ucl.ac.uk/1419156/.
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Potential therapeutic avenues may emerge from understanding morphological adaptations of adult human skeletal muscle that result from critical issue ischaemia (CLI). There is little understanding of the role and limitations of satellite cells in repairing and regenerating ischaemic tissue. This study aims to show the pathognomonic changes that take place as adaptations to chronic ischaemia. Patients (n=10) undergoing lower limb amputations for CLI were recruited to the study and gastrocnemius muscle biopsies were compared to those from control patients (n=10) undergoing long saphenous vein harvesting for coronary artery bypass grafting. Transmission Electron microscopy, histology, immunohistochemistry and Western blotting were used to assess the myogenic response to ischaemia. Any significant change in tissue morphology, morphometry and satellite cell number or activity was of interest. There was significantly greater deposition of fibrofatty tissue, collagen and a loss of polygonal structure in CLI samples. Myonuclear number per fibre was not significantly different, neither was the occurrence of centrally occurring nuclei. All fibre types demonstrated significant atrophy except Ilc. Ilc hybrid fibres were more abundant in CLI samples (p=0.0147). Type I fibres displayed a proportionate rise by 2.4 fold (p=0.0288). Type Ilx fibres were most susceptible to ischaemia with x6 fold reduction in number (p=0.0039) and greatest reduction in CSA (p=0.0029). Type II fibres showed greater fibre-size diversity. The endothelial marker, CD31 and the haematopoietic stem cell marker, CD34 were more abundant in CLI (p<0.0001). There was over expression of the satellite cell marker pax? (p<0.0001) and quiescent satellite cell numbers. MyoD, a marker of activated satellite cells is significantly reduced in ischaemia (p<0.0001). These findings confirm active repair and regeneration in CLI tissue, but the indigenous response of muscle is inadequate for proper healing. These processes are disordered, with limited maturation of myogenesis.
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Guest, Kay P. "Mathematical modelling of the half-sarcomere from a human skeletal muscle." Thesis, University of Warwick, 2013. http://wrap.warwick.ac.uk/60665/.
Full textAbstract:
The character of the functional output of a motor unit within skeletal muscle has been linked experimentally to the proteins found in the sarcomere, the smallest contractile unit of muscle fibre. Current mathematical models focus on either individual chemical reactions or the bulk properties of muscle with limited reference to the internal processes and structures within the muscle. Without an understanding of those internal properties, the normal function of muscle cannot be simulated and consequently muscular diseases and their treatments cannot be accurately modelled. In this project, a mathematical model has been developed which relates the chemomechanical cycle of individual events (crossbridges) to the transfer of mechanical energy through an actin filament, myosin cofilament and, by incorporating the protein titin, the mechanical properties of the interconnecting proteins in a section of sarcomere. Evaluation and parameterisation of the model were made by comparison with in vitro test data from the published literature at the level of a single crossbridge and single filaments. At the single filament level, the model was evaluated against two conditions: a low load high displacement (concentric contraction) and a high load low displacement (isometric contraction). In isometric loading the peak force level per unit length of actin filament was higher than that observed in vitro, the difference being attributable to the greater compliance in the substrate used in vitro to hold the myosin fragments (~37pN compared to ~12pN). The mean number of concurrent crossbridges was consistent between the model and in vitro data. Under low load the model demonstrated filament movement at speeds comparable to those measured in in vitro motility studies, although longer filaments in the model were required than those in vitro to reach the higher speeds (7μm vs. 2μm for ~8μm/s). By making the pre-lever reaction duration of the crossbridge cycle strain dependent it was possible to obtain long reaction cycles in low load scenarios comparable to those observed for fragments in solution while generating the actin filament speeds observed in vitro. It was necessary to have a distribution of attachment times across the filament in order to generate and maintain filament movement in the model; the variation being governed by the tension distribution in the filament. By applying a passive loading as generated by the titin protein the filaments moved more rapidly, with an increased contribution from each crossbridge to filament movement. Initial results indicate examination of the strain dependency of the post-lever reaction duration may modify filament speeds and will increase the proportion of each crossbridge movement that contributes to the actin filament propulsion (increase crossbridge efficiency). Examination of a selection of the model’s parameters gave an initial evaluation of how the model could be ‘tuned’ to change the number, reaction state and distribution in time of crossbridges to achieve changes in filament contraction speed, isometric force generation and the efficiency with which crossbridges are used; noting that one desired output may conflict with another. The interaction of the passive components in the structure of the sarcomere with the strain dependent reaction cycle at each crossbridge demonstrated the potential limitations of scaling and averaging localised events without consideration of the passive structures present in the fibre and muscle bulk. The model provides a means to examine the mechanisms and parameters of the sarcomere’s function and how those parameters may be adjusted to achieve different output characteristics. The model provides a foundation for the emulation of muscle fibres and a motor unit in health and disease.